Potent and safe inhibitors of anthrax lethal factor Anthrax is primarily a disease of herbivores caused by gram-positive, aerobic, spore-forming B.anthracis^ Humans are accidental hosts and cases of anthrax have been reported from almost every country. The existence of antibiotic:resistant bacterial strains that arise either naturally or through deliberate engineering further emphasizes the need for alternative therapeutic approaches. Vaccines are typically problematic for prophylactic treatment of large civilian groups, because the inevitable side-effects must somehow be weighed against the chances of epidemics. The progression of anthrax depends on a secreted toxin, LF (lethal factor), a protease that plays a key role in inducing apoptosis of macrophages by cleaving the specific human cell signaling proteins. Anti-toxin approaches that treat infection therapeutically by targeting LF should therefore provide a promising alternative or complement to antibiotics and vaccines. However, further research on the role of LF for the development of Anthrax therapies is hampered by the lack of potent and selective pharmacological tools. Based on these observations, a central hypothesis in our research proposal is that small organic molecules capable of blocking or reducing the activity of LF would render B. Anthracis avirulent, making Anthrax disease treatable in combination with conventional antibiotics. We propose here to develop a novel series of LF inhibitors based on a multidisciplinary approach that involves fragment-based lead design, medicinal chemistry and iterative .structure-based optimizations. In vitro and finally in vivo efficacy studies will be conducted on the most promising agents. Our three specific aims are: (I) To accomplish structure-based design of potent inhibitors capable of selectively blocking the LF cleavage activity without affecting human MMP enzymes. (II) To increase, through derivatizations and iterations, the selectivity, potency and safety of the selected inhibitor scaffolds. (Ill) To test the effectiveness of fine-tuned compounds in protecting the cultured cells and to identify the drug-like lead inhibitors) of LF for continued drug refinement, and in.vivo efficacy studies. Relevance Inhaling B.anthracis spores causes inhalation anthrax, the most deadly form of the disease. As the disease progresses, the spores, engulfed by alveolar macrophages, germinate to generate the bacteria, which spread through the lymph nodes to the bloodstream, eventually leading to systemic fatal shock. To successfully accomplish invasion, B.anthracis has to weaken the host immune system. LF (lethal factor) is a secreted toxin that plays a key role in inducing apoptosis (cell-death) of macrophages by cleaving the specific human cell signaling proteins. Conversely, specific inhibition of LF should prevent macrophage cell- death and promote termination of the disease. The inhibitors of LF we will develop, most likely when used in combination with antibiotics, should facilitate and assure the recovery of patients who harbor the disease.